TDR Targets ID: 4387 P. falciparum 3D7, dihydroorotate dehydrogenase, mitochondrial precursor http://www.tdrtargets.org/targets/view?gene_id=4387 *Target (protein/gene name): dihydroorotate dehydrogenase *NCBI Gene # 3885966 or RefSeq#: NC_004327.2 *Protein ID (NP or XP #) or Wolbachia#: XP_966023.1 *Organism (including strain):Plasmodium falciparum 3D7 Etiologic Risk Group (see link below):Appendix B-II-C. Risk Group 2 (RG2) - Parasitic Agents *Background/Disease Information (sort of like the Intro to your Mini Research Write up): Plasmodium falciparum is a protozoan parasite that can cause malaria in humans. Female mosquitos transmit it. Malaria can be transmitted by other organisms, but ¾ of the cases are caused by plasmodium falciparum. This is also the most deadly of all malaria. Essentiality of this protein:Phenotype: significant loss of fitness in bloodstream forms (6 days); Source study: alsford; http://www.tdrtargets.org/targets/view?gene_id=4387
"Analysis of bloodstream and insect life-cycle stages and differentiated libraries revealed genome-scale knockdown profiles of growth and development, linking thousands of previously uncharacterized and "hypothetical" genes to essential functions. Genes underlying prominent features of trypanosome biology are highlighted, including the constitutive emphasis on post-transcriptional gene expression control, the importance of flagellar motility and glycolysis in the bloodstream, and of carboxylic acid metabolism and phosphorylation during differentiation from the bloodstream to the insect stage." http://www.ncbi.nlm.nih.gov/pubmed/21363968
"Pyrimidine biosynthesis presents an attractive drug target in malaria parasites due to the absence of a pyrimidine salvage pathway. A set of compounds designed to inhibit the Plasmodium falciparum pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (PfDHODH) was synthesized." http://pubchem.ncbi.nlm.nih.gov/assay/assay.cgi?aid=275413&loc=ea_ras#aDescription
Structure Available (PDB or Homology model) http://www.pdb.org/pdb/explore/explore.do?structureId=1TV5 -- PDB # or closest PDB entry if using homology model: -- For Homology Model option: ---- Show pairwise alignment of your BLASTP search in NCBI against the PDB ---- Query Coverage: ---- Max % Identities: ---- % Positives ---- Chain used for homology: Current Inhibitors: "Both teriflunomide and leflunomide are inhibitors of the mitochondrial enzyme dihydroorotate dehydrogenase, which is critically involved in pyrimidine synthesis." http://www.ncbi.nlm.nih.gov/pubmed/21157651
Expression Information (has it been expressed in bacterial cells): From Brenda information, this organism has been expressed in Lactobacillus delbrueckii subsp. bulgaricus, Crithidia fasciculata, Escherichia coli, Lactococcus lactis, Homo sapiens, Sulfolobus solfataricus, and Trypanosoma cruzi
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'): misklkpqfm flpkkhilsy crkdvlnlfe qkfyytskrk esnnmknesl lrlinynryy nkidsnnyyn ggkilsndrq yiysplceyk kkindissyv svpfkinirn lgtsnfvnnk kdvldndyiy enikkekskh kkiifllfvs lfglygffes ynpefflydi flkfclkyid geichdlfll lgkynilpyd tsndsiyact nikhldfinp fgvaagfdkn gvcidsilkl gfsfieigti tprgqtgnak prifrdvesr siinscgfnn mgcdkvtenl ilfrkrqeed kllskhivgv sigknkdtvn ivddlkycin kigryadyia invsspntpg lrdnqeagkl kniilsvkee idnleknnim ndestynedn kivekknnfn knnshmmkda kdnflwfntt kkkplvfvkl apdlnqeqkk eiadvlletn idgmiisntt tqindiksfe nkkggvsgak lkdistkfic emynytnkqi piiasggifs gldalekiea gasvcqlysc lvfngmksav
qikrelnhll yqrgyynlke aigrkhsks *length of your protein in Amino Acids: 569 Molecular Weight of your protein in kiloDaltons using the**Expasy ProtParam** website: 65558.4 Molar Extinction coefficient of your protein at 280 nm wavelength:51690 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. *CDS Gene Sequence (paste as text only):ATGATCTCTAAATTGAAACCTCAATTTATGTTTTTACCAAAGAAACATATTTTAAGTTATTGTAGAAAGGATGTTTTAAATTTGTTTGAACAGAAGTTTTATTATACTAGCAAACGGAAAGAAAGTAATAATATGAAGAATGAATCTTTATTAAGATTAATTAATTATAATAGATATTATAATAAGATAGATTCTAATAATTATTATAATGGTGGAAAAATATTAAGTAATGATAGGCAATATATATATTCACCATTATGTGAATATAAAAAGAAAATAAATGATATATCATCATATGTATCTGTACCTTTTAAGATTAATATAAGAAATTTAGGTACTTCCAATTTTGTAAATAATAAGAAGGATGTACTTGATAATGATTATATTTATGAAAATATTAAAAAAGAAAAATCTAAGCATAAAAAAATAATATTTTTATTATTTGTTTCATTATTTGGATTATATGGTTTTTTTGAATCTTATAATCCTGAATTTTTTTTATATGATATATTTTTAAAATTCTGTTTAAAATATATTGATGGTGAAATATGTCATGACCTTTTTTTATTACTAGGAAAATATAATATATTACCATATGATACTAGTAATGATAGTATATATGCATGTACAAATATTAAACATCTTGATTTTATAAATCCATTCGGTGTTGCTGCAGGATTTGATAAAAACGGTGTATGTATAGATAGCATATTAAAATTAGGGTTTTCGTTTATCGAAATTGGTACCATAACCCCAAGGGGCCAAACGGGTAATGCAAAACCACGTATTTTTAGAGACGTTGAATCTAGAAGTATTATAAATTCATGTGGCTTTAATAATATGGGTTGTGACAAAGTTACAGAAAATTTAATACTTTTTCGTAAAAGACAAGAAGAAGATAAATTGTTAAGTAAACATATTGTAGGTGTCAGTATAGGTAAGAATAAAGATACTGTTAATATTGTAGATGATCTAAAATATTGTATTAATAAAATAGGAAGATACGCTGATTATATAGCTATTAATGTAAGCTCCCCTAATACACCTGGGTTAAGAGATAATCAAGAAGCTGGGAAGTTAAAAAATATAATTTTAAGTGTAAAAGAAGAAATAGATAATTTAGAAAAGAATAATATTATGAATGATGAAAGTACTTATAATGAAGATAATAAAATAGTAGAAAAAAAAAATAATTTTAATAAAAATAATAGTCACATGATGAAAGATGCTAAGGATAACTTCTTATGGTTTAATACAACAAAAAAGAAGCCCTTGGTTTTTGTTAAGTTAGCTCCAGATCTTAATCAAGAACAGAAAAAAGAAATTGCTGATGTATTACTTGAAACTAATATAGATGGTATGATTATTTCTAATACTACGACACAAATAAATGACATAAAAAGTTTTGAAAATAAAAAAGGAGGTGTTAGCGGAGCAAAACTAAAAGATATATCTACAAAATTTATATGTGAAATGTATAATTATACAAATAAACAAATACCCATTATTGCATCAGGAGGGATATTTAGTGGATTGGATGCTTTAGAAAAAATTGAAGCAGGTGCTTCAGTTTGTCAATTATATTCTTGTTTGGTTTTTAATGGTATGAAATCAGCTGTACAAATAAAAAGAGAATTGAATCACTTGCTATATCAAAGAGGATATTACAATTTAAAGGAGGCCATTGGCCGAAAGCATAGCAAAAGTTAA *GC% Content for gene: 24.561403508772% *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): *GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now) Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers): (link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol) -- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
P. falciparum 3D7, dihydroorotate dehydrogenase, mitochondrial precursor
http://www.tdrtargets.org/targets/view?gene_id=4387
*Target (protein/gene name): dihydroorotate dehydrogenase
*NCBI Gene # 3885966 or RefSeq#: NC_004327.2
*Protein ID (NP or XP #) or Wolbachia#: XP_966023.1
*Organism (including strain): Plasmodium falciparum 3D7
Etiologic Risk Group (see link below):Appendix B-II-C. Risk Group 2 (RG2) - Parasitic Agents
*Background/Disease Information (sort of like the Intro to your Mini Research Write up):
Plasmodium falciparum is a protozoan parasite that can cause malaria in humans. Female mosquitos transmit it. Malaria can be transmitted by other organisms, but ¾ of the cases are caused by plasmodium falciparum. This is also the most deadly of all malaria.
Essentiality of this protein: Phenotype: significant loss of fitness in bloodstream forms (6 days); Source study: alsford; http://www.tdrtargets.org/targets/view?gene_id=4387
"Analysis of bloodstream and insect life-cycle stages and differentiated libraries revealed genome-scale knockdown profiles of growth and development, linking thousands of previously uncharacterized and "hypothetical" genes to essential functions. Genes underlying prominent features of trypanosome biology are highlighted, including the constitutive emphasis on post-transcriptional gene expression control, the importance of flagellar motility and glycolysis in the bloodstream, and of carboxylic acid metabolism and phosphorylation during differentiation from the bloodstream to the insect stage."
http://www.ncbi.nlm.nih.gov/pubmed/21363968
"Pyrimidine biosynthesis presents an attractive drug target in malaria parasites due to the absence of a pyrimidine salvage pathway. A set of compounds designed to inhibit the Plasmodium falciparum pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (PfDHODH) was synthesized."
http://pubchem.ncbi.nlm.nih.gov/assay/assay.cgi?aid=275413&loc=ea_ras#aDescription
Complex of proteins?: no
Druggable Target: yes
*EC#: 1.3.98.1
Link to BRENDA EC# page:
http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.3.98.1
-- Show screenshot of BRENDA enzyme mechanism schematic
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
"Dihydroorotate oxidation was usually measured by following ferricyanide reduction (reaction 1, below) as previously described (15). Where indicated, 2, 6-dichlorophenolindolphenol 98 (DCI) (0.2 usmol) or cytochrome c (1 mg) was substituted for ferricyanide in the standard protocol. Reduction of the acceptor was followed spectrophotometrically at 600 nm for DCI and at 550 nm for cytochrome c as described previously."
-- link to Sigma (or other company) page for assay or assay reagents (substrates)
http://jb.asm.org/content/119/1/98.long
-- link (or citation) to paper that contains assay information
http://pubchem.ncbi.nlm.nih.gov/assay/assay.cgi?aid=275413&loc=ea_ras
-- List cost and quantity of substrate reagents and supplier
http://www.sigmaaldrich.com/catalog/search?interface=All&term=fumarate&lang=en®ion=US&focus=product&N=0+220003048+219853269+219853286
http://www.molport.com/buy-chemicals/moleculelink/4S-2-6-dioxo-1-3-diazinane-4-carboxylic-acid/3941134
Structure Available (PDB or Homology model)
http://www.pdb.org/pdb/explore/explore.do?structureId=1TV5
-- PDB # or closest PDB entry if using homology model:
-- For Homology Model option:
---- Show pairwise alignment of your BLASTP search in NCBI against the PDB
---- Query Coverage:
---- Max % Identities:
---- % Positives
---- Chain used for homology:
Current Inhibitors:
"Both teriflunomide and leflunomide are inhibitors of the mitochondrial enzyme dihydroorotate dehydrogenase, which is critically involved in pyrimidine synthesis."
http://www.ncbi.nlm.nih.gov/pubmed/21157651
Expression Information (has it been expressed in bacterial cells):
From Brenda information, this organism has been expressed in Lactobacillus delbrueckii subsp. bulgaricus, Crithidia fasciculata, Escherichia coli, Lactococcus lactis, Homo sapiens, Sulfolobus solfataricus, and Trypanosoma cruzi
Purification Method: http://www.ncbi.nlm.nih.gov/pubmed/7727509
Image of protein (PyMol with features delineated and shown separately):
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
misklkpqfm flpkkhilsy crkdvlnlfe qkfyytskrk esnnmknesl lrlinynryy
nkidsnnyyn ggkilsndrq yiysplceyk kkindissyv svpfkinirn lgtsnfvnnk
kdvldndyiy enikkekskh kkiifllfvs lfglygffes ynpefflydi flkfclkyid
geichdlfll lgkynilpyd tsndsiyact nikhldfinp fgvaagfdkn gvcidsilkl
gfsfieigti tprgqtgnak prifrdvesr siinscgfnn mgcdkvtenl ilfrkrqeed
kllskhivgv sigknkdtvn ivddlkycin kigryadyia invsspntpg lrdnqeagkl
kniilsvkee idnleknnim ndestynedn kivekknnfn knnshmmkda kdnflwfntt
kkkplvfvkl apdlnqeqkk eiadvlletn idgmiisntt tqindiksfe nkkggvsgak
lkdistkfic emynytnkqi piiasggifs gldalekiea gasvcqlysc lvfngmksav
qikrelnhll yqrgyynlke aigrkhsks
*length of your protein in Amino Acids: 569
Molecular Weight of your protein in kiloDaltons using the **Expasy ProtParam** website: 65558.4
Molar Extinction coefficient of your protein at 280 nm wavelength: 51690
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
*CDS Gene Sequence (paste as text only):ATGATCTCTAAATTGAAACCTCAATTTATGTTTTTACCAAAGAAACATATTTTAAGTTATTGTAGAAAGGATGTTTTAAATTTGTTTGAACAGAAGTTTTATTATACTAGCAAACGGAAAGAAAGTAATAATATGAAGAATGAATCTTTATTAAGATTAATTAATTATAATAGATATTATAATAAGATAGATTCTAATAATTATTATAATGGTGGAAAAATATTAAGTAATGATAGGCAATATATATATTCACCATTATGTGAATATAAAAAGAAAATAAATGATATATCATCATATGTATCTGTACCTTTTAAGATTAATATAAGAAATTTAGGTACTTCCAATTTTGTAAATAATAAGAAGGATGTACTTGATAATGATTATATTTATGAAAATATTAAAAAAGAAAAATCTAAGCATAAAAAAATAATATTTTTATTATTTGTTTCATTATTTGGATTATATGGTTTTTTTGAATCTTATAATCCTGAATTTTTTTTATATGATATATTTTTAAAATTCTGTTTAAAATATATTGATGGTGAAATATGTCATGACCTTTTTTTATTACTAGGAAAATATAATATATTACCATATGATACTAGTAATGATAGTATATATGCATGTACAAATATTAAACATCTTGATTTTATAAATCCATTCGGTGTTGCTGCAGGATTTGATAAAAACGGTGTATGTATAGATAGCATATTAAAATTAGGGTTTTCGTTTATCGAAATTGGTACCATAACCCCAAGGGGCCAAACGGGTAATGCAAAACCACGTATTTTTAGAGACGTTGAATCTAGAAGTATTATAAATTCATGTGGCTTTAATAATATGGGTTGTGACAAAGTTACAGAAAATTTAATACTTTTTCGTAAAAGACAAGAAGAAGATAAATTGTTAAGTAAACATATTGTAGGTGTCAGTATAGGTAAGAATAAAGATACTGTTAATATTGTAGATGATCTAAAATATTGTATTAATAAAATAGGAAGATACGCTGATTATATAGCTATTAATGTAAGCTCCCCTAATACACCTGGGTTAAGAGATAATCAAGAAGCTGGGAAGTTAAAAAATATAATTTTAAGTGTAAAAGAAGAAATAGATAATTTAGAAAAGAATAATATTATGAATGATGAAAGTACTTATAATGAAGATAATAAAATAGTAGAAAAAAAAAATAATTTTAATAAAAATAATAGTCACATGATGAAAGATGCTAAGGATAACTTCTTATGGTTTAATACAACAAAAAAGAAGCCCTTGGTTTTTGTTAAGTTAGCTCCAGATCTTAATCAAGAACAGAAAAAAGAAATTGCTGATGTATTACTTGAAACTAATATAGATGGTATGATTATTTCTAATACTACGACACAAATAAATGACATAAAAAGTTTTGAAAATAAAAAAGGAGGTGTTAGCGGAGCAAAACTAAAAGATATATCTACAAAATTTATATGTGAAATGTATAATTATACAAATAAACAAATACCCATTATTGCATCAGGAGGGATATTTAGTGGATTGGATGCTTTAGAAAAAATTGAAGCAGGTGCTTCAGTTTGTCAATTATATTCTTGTTTGGTTTTTAATGGTATGAAATCAGCTGTACAAATAAAAAGAGAATTGAATCACTTGCTATATCAAAGAGGATATTACAATTTAAAGGAGGCCATTGGCCGAAAGCATAGCAAAAGTTAA
*GC% Content for gene: 24.561403508772%
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only):
*GC% Content for gene (codon optimized):
Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
Resources:
http://www.tdrtargets.org/targets/view?gene_id=4387
http://www.ncbi.nlm.nih.gov/gene/?term=dihydroorotate+dehydrogenase+falciparum
http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.3.98.1
http://www.brenda-enzymes.org/literature/lit.php4?e=1.3.98.1&r=390909
http://www.brenda-enzymes.org/literature/lit.php4?e=1.3.98.1&r=390909
http://en.wikipedia.org/wiki/Dihydroorotate_dehydrogenase
http://www.rcsb.org/pdb/results/results.do?outformat=&qrid=8F410442&tabtoshow=Current
http://www.bindingdb.org/bind/ByMonomersTarget.jsp
http://www.ncbi.nlm.nih.gov/gene/?term=dihydroorotate+dehydrogenase
http://jb.asm.org/content/119/1/98.long